DE69606552T2 - ACYLATED ENOL DERIVATIVES OF ALPHA KETOESTERS AND ALPHA KETOAMIDES - Google Patents

Description

BACKGROUND OF THE INVENTION

This invention relates to acylated enol derivatives of α-ketoesters and α-ketoamides, which exist either as elastase inhibitors or prodrugs of elastase inhibitors and are useful for a variety of physiological and end-use applications.

Human neutrophil elastase has been implicated as an agent contributing to the tissue destruction associated with a number of inflammatory diseases such as chronic bronchitis, cystic fibrosis, and rheumatoid arthritis (J. L. Melech and J. I. Gallin, New Engl. J. Med. 317 (11) (1987), 687). Elastase exhibits a broad range of proteolytic activity against a number of connective tissue macromolecules, including elastin, fibronectin, collagen, and proteoglycan. The presence of the enzyme elastase may contribute to the pathology of these diseases.

Normal plasma contains large amounts of protease inhibitors, which control a number of enzymes involved in the turnover and inflammation of connective tissue. For example, α-1-proteinase inhibitor (α-1-PI) is a serine protease inhibitor that blocks elastase activity. α-1-PI has received significant interest because reductions in plasma levels to less than 15% of normal have been associated with the early development of emphysema.

In addition to plasma-derived protease inhibitors, secretions, including bronchial, nasal, cervical mucus, and the Seminal fluid contains an endogenous protease inhibitor, called secretory leukoprotease inhibitor (SLPI), which can inactivate elastase and is thought to play an important role in maintaining the integrity of the epithelium in the presence of proteases from inflammatory cells. In certain pathological states, α-1-PI and SLPI are inactivated by neutrophil oxidative mechanisms, allowing neutrophil proteases to function in an essentially inhibitor-free environment. For example, bronchial lavage fluids from patients with respiratory distress syndrome (ARDS) have been found to contain active elastase and α-1-PI, which have been inactivated by oxidation.

In addition to oxidative mechanisms, neutrophils have non-oxidative mechanisms to evade inhibition by antiproteases. Neutrophils from patients with chronic septic granulomatosis can deplete endothelial precursors in the presence of excess α-1-PI. In vitro, there is considerable evidence that stimulated neutrophils can bind so tightly to their substrates that serum antiproteases are effectively excluded from the microenvironment of intimate cell-substrate contact. The influx of large numbers of neutrophils to an inflammatory site can result in significant tissue destruction due to the proteolysis that occurs in this region.

Applicants have previously determined that elastase is one of the primary neutrophil proteases responsible for cartilage matrix degeneration, as measured by the ability of neutrophil lysate, purified elastase and stimulated neutrophils to degrade cartilage matrix proteoglycan. Furthermore, various types of peptide derivatives are known in the art which are useful as elastase inhibitors and exhibit valuable pharmacological activities. For example, Angelastro, M. R. et al., J. Med. Chem. 37 (1994), 4538 and European Patent Application OPI No. 0529568, inventors Peet et al., with a publication date of March 3, 1993, disclose that various peptides, such as valylprolylvalylpentafluoroethyl ketones, with different N-protecting groups are inhibitors of human neutrophil elastase (HNE) in vitro and in vivo and are also orally active in models of HNE-induced rat and hamster pulmonary hemorrhage.

In addition, the art discloses that a number of different amino acid moieties are permissible at the P2, P3 and P4 sites of the elastase inhibitor and that a number of the N-protecting groups can be substituted while still retaining enzyme inhibitory activity, although oral activity is only noted for some N-protecting groups. For example, Skiles, J. W. et al., J. Med. Chem. 35 (1992), 641 discloses a number of tripeptide elastase inhibitors which have trifluoromethyl groups or aryl ketone residues at P1 and N-substituted glycine residues at P2. It is shown that no dramatic change in in vitro potency is observed when the substituent on P2-glycine is increased in size and lipophilicity in the range of H, CH3, cyclopentyl, exonorbornyl, 2-indanyl, cycloheptyl, cyclooctyl and also piperidinyl, benzyl, 3,4-dimethoxyphenethyl, tetrahydrofurfuryl and furfuryl.

Similarly, U.S. Patent No. 4,910,190, issued March 20, 1990 to Bergeson et al., and U.S. Pat. No. 5,194,588, issued March 16, 1993 to Edwards et al., and published European Patent Application No. 0195212, inventor Michael Kolb et al., published September 24, 1986, teach that a number of alkyl and substituted alkyl radicals are permissible as side chain groups of the amino acids at the P1 and P4 positions. In addition, elastase inhibitors containing typical N-protecting groups such as acetyl, succinyl, t-butyloxycarbonyl, carbobenzyloxy, 4-((4-chlorophenyl)sulfonylaminocarbonyl)phenylcarbonyl groups and the like were specifically disclosed.

As prodrugs of elastase inhibitors, various analogous compounds of N-[4-(4-morpholinylcarbonyl)benzoyl]-L-valyl-N-[3,3,4,4,4-pentafluoro-1-(1-methylethyl)-2-oxobutyl]-L-prolinamide, in which the chiral center of the residue P1 has been eliminated, are known by Burkhart, J. P. et al., J. Med. Chem. 38 (1995), 223. Applicants recently discovered acetylated enol derivatives of known non-fluorinated elastase inhibitors, which are useful as prodrugs of elastase inhibitors or are themselves elastase inhibitors.

SUMMARY OF THE INVENTION

The present invention relates to compounds of the formulas

or

where

R₁ is a (C₁-C₄)alkyl radical,

R₂ is a (C₁-C₄)alkyl radical, a phenyl, benzyl, cyclohexyl or cyclohexylmethyl group,

X is -CO₂R₃ or -CONHR₃', where R₃ is a (C₁-C₄)alkyl radical, a phenyl, benzyl, cyclohexyl or cyclohexylmethyl group and R₃' is a hydrogen atom, a (C₁-C₄)alkyl radical, a phenyl, benzyl, cyclohexyl or cyclohexylmethyl group,

P₂ is Gly or Ala, wherein the nitrogen of the α-amino group is optionally substituted with a radical R, wherein R is (C₁-C₆)alkyl, (C₃-C₁₂)cycloalkyl, (C₃-C₁₂)cycloalkyl-(C₁-C₆)alkyl, (C₄-C₁₁)bicycloalkyl, (C₄-C₁₁)bicycloalkyl-(C₁-C₆)alkyl, (C₆-C₁₁)aryl, (C₆-C₁₀)aryl-(C₁-C₆)alkyl, (C3 -C7 )heterocycloalkyl, (C3 -C7 )heterocycloalkyl-(C1 -C6 )alkyl, (C5 -C9 )heteroaryl, (C5 -C9 )heteroaryl(C1 -C6 )alkyl, fused (C6 -C10 )-aryl-(C3 -C12 )cycloalkyl, fused (C6 -C10 )aryl-(C3 -C12 )cycloalkyl-(C1 -C6 )alkyl, fused (C5 -C9 )heteroaryl-(C3 -C12 )cycloalkyl or fused (C5 -C9 )heteroaryl(C3 -C12 )cycloalkyl(C1 -C6 -)alkyl, or P&sub2; Pro, Aze, Ind, Tic, Pip, Tca, Pro(4-OBzl), Pro(4-OAc), Pro(4-OH) is,

P₃ is Ala, bAla, Leu, Ile, Nle, Val, Nva, Lys or bVal,

P₄ Ala, bAla, Val, Nva, bVal, Pro or absent,

K is a hydrogen atom, an acetyl, succinyl, benzoyl, t-butyloxycarbonyl, carbobenzyloxy, dansyl, isovaleryl, methoxysuccinyl, 1-adamantanesulfonyl, 1-adamantaneacetyl, 2-carboxybenzoyl group, -C(O) N(CH3 )2 , 4-((chlorophenyl)sulfonylaminocarbonyl)phenylcarbonyl, 4-((4-bromophenyl)sulfonylaminocarbonyl)phenylcarbonyl, 4-(sulfonylaminocarbonyl)phenylcarbonyl or a residue of the formulas

or

where Z is N or CH, B is a radical of the formulas

or

(where the wavy line represents the binding site to the rest of the molecule, i.e. not to Z),

R' is a hydrogen atom or a (C₁-C₄)alkyl radical,

n is the number 0 or the integer 1 or 2,

or a hydrate or a pharmaceutically acceptable salt thereof, which are useful as prodrugs of known elastase inhibitors or inhibit elastase in its present form. The compounds of formulas I or IA thus either exhibit an anti-inflammatory activity which is useful in the treatment of emphysema, cystic fibrosis, shock lung, septicemia, disseminated intravascular coagulation, gout, rheumatoid arthritis, chronic bronchitis and inflammatory irritable bowel syndrome, or they are prodrugs of compounds which exhibit such effects.

DETAILED DESCRIPTION OF THE INVENTION

As used herein, the term "(C₁-C₄)alkyl" means a straight or branched chain alkyl group having 1 to 4 carbon atoms, such as methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl and the like. Likewise, the term "(C₁-C₆)alkyl" means a straight or branched chain alkyl group having 1 to 6 carbon atoms, such as methyl, ethyl, n-propyl, isopropyl, n-butyl, n-pentyl, sec-pentyl, isopentyl and n-hexyl. The term "(C₃-C₁₂)cycloalkyl" means a cyclic alkyl group consisting of a 3- to 12-membered ring which may be substituted by a lower alkyl group, for example a cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, 4-methylcyclohexyl, 4-ethylcyclohexyl, cycloheptyl and cyclooctyl group. The term "(C₃-C₁₂)cycloalkyl-(C₁-C₆)alkyl" means a (C₁-C₆)alkyl group which is substituted by a (C₃-C₁₂)cycloalkyl group, such as a cyclohexylmethyl or cyclopentylethyl group. The term "(C₄-C₁₁)bicycloalkyl" means an alkyl group containing a pair of bridging carbon atoms, such as 2-bicyclo[1.1.0]butyl, 2-bicyclo[2.2.1]hexyl and 1-bicyclo[2.2.2]octane. The term "(C₄-C₁₁)bicycloalkyl-(C₁-C₆)alkyl" means a (C₁-C₆)alkyl group substituted by a (C₆-C₁₁)bicycloalkyl group, such as 2-bicyclohexylmethyl. The term "(C₆-C₁₀)-aryl group means a cyclic aromatic combination of conjugated carbon atoms, e.g. a phenyl, 1-naphthyl and 2-naphthyl group. The term "(C₆-C₁₀)-aryl-(C₁-C₆)-alkyl group" means a (C₁-C₆)-alkyl group which is substituted by a (C₆-C₁₀)-aryl group, such as a benzyl, phenethyl and 1-naphthylmethyl group. The term "(C₃-C₇)-heterocycloalkyl group" means a non-aromatic carbon containing cyclic radical containing 1 to 3 heteroatoms selected from oxygen, nitrogen and sulfur, such as morphinyl and piperidinyl. The term "(C₃-C₇)heterocycloalkyl-(C₁-C₆)alkyl" means a (C₁-C₆)alkyl radical substituted by a (C₃-C₇)heterocycloalkyl radical, e.g. morpholinomethyl. The term "(C₅-C₆)heteroaryl radical" means a cyclic or bicyclic aromatic combination of conjugated carbon atoms and 1 to 3 nitrogen, oxygen and sulfur atoms, e.g. pyridinyl, 2-quinoxalinyl and quinolinyl. The term "(C₅-C₉)heteroaryl-(C₁-C₆)alkyl" means a (C₁-C₆)alkyl which is substituted by a (C₅-C₉)heteroaryl group, such as a 3-quinolinylmethyl group. The term "fused (C₆-C₁₀)aryl-(C₃-C₁₂)cycloalkyl" means a (C₃-C₁₂)cycloalkyl group having one or more side chains shared with a (C₆-C₁₀)aryl group and which may include groups obtained, for example, by the condensation of benzene and cyclopentane, such as a 2-indanyl group. The term "fused (C6-C10)aryl-(C3-C12)cycloalkyl-(C1-C6)alkyl" means a (C1-C6)alkyl radical which is substituted by a fused (C6-C10)aryl-(C3-C12)cycloalkyl radical. The term "fused (C5-C9)heteroaryl-(C3-C8)cycloalkyl radical" means a (C5-C9)heteroaryl radical having one or more side chains shared with a (C3-C8)cycloalkyl radical, which may be, for example, B. can include groups obtained by the condensation of cyclohexane and pyridine, i.e. a tetrahydroquinoline group. Finally, the term "condensed (C₅-C₉)-heteroaryl-(C₃-C₈)-cycloalkyl-(C₁-C₆)-alkyl radical" means a (C₁-C₆)-alkyl radical which is substituted by a condensed (C₅-C₉)-heteroaryl-(C₃-C₈)-cycloalkyl radical.

The term "stereoisomers" is a general term for all isomers of individual molecules that differ only in the orientation of their atoms in space. It includes mirror image isomers (enantiomers), geometric (cis/trans) isomers, and isomers of compounds with more than one chiral center that are not mirror images of another (diastereomers). For amino acids, the designations L/D or R/S can be used, as described in IUPAC-IUB Joint Commission on Biochemical Nomenclature, Eur. J. Biochem. 138 (1984), 9-37.

The term "pharmaceutically acceptable salt" refers to those salts which, at the administered dose, are essentially insufficient to achieve the desired effect. toxic and do not independently possess significant pharmacological activity. The salts included within the scope of this term are salts with hydrobromic, hydrochloric, sulphuric, phosphoric, nitric, formic, acetic, propionic, succinic, glycolic, lactic, malic, tartaric, citric, ascorbic, α-ketoglutaric, glutamic, aspartic, maleic, hydroxymaleic, pyruvic, phenylacetic, benzoic, p-aminobenzoic, anthranilic, p-hydroxybenzoic, salicylic, hydroxyethanesulphone, ethylenesulphone, halobenzenesulphone, toluenesulphone, naphthalenesulphone, methanesulphone, sulphanilic acid and the like.

Each α-amino acid has a distinctive "R residue," where R is the side chain or residue attached to the α-carbon of the α-amino acid. For example, the R side chain for glycine is a hydrogen atom, for alanine a methyl group, and for valine an isopropyl group. For the specific R residues or side chains of α-amino acids, see A. L. Lehninger's text on Biochemistry.

Unless otherwise stated, the α-amino acids of these peptidase substrate analogs are preferably in their L-configuration, however, the authors contemplate that the amino acids of the compounds of formula I or IA may exist as either D- or L-configurations or as mixtures of D- and L-isomers, including the racemic mixture. The accepted abbreviations for the amino acids are set forth in Table I.

Table I Amino acid symbol

Alanine Ala

Glycine Gly

Isoleucine Ile

Leucine Leu

Lysine Lys

Phenylalanine Phe

Proline Pro

Tryptophan Trp

Tyrosine Tyr

Valine Val

Norvalin Nva

Norleucine Nle

2-Indolinecarboxylic acid Ind

Beta-Alanine BAla

Methionine Met

Azetidinecarboxylic acid Aze

4-Acetoxyproline Pro(4-OAc)

4-Benzyloxyproline Pro(4-OBzl)

4-Hydroxyproline Pro(4-OH)

Pipecolic acid Pip

Thiazolidine-4-carboxylic acid Tca

1,2,3,4-Tetrahydro-3-isoquinolinecarboxylic acid Tic

Beta-Valine BVal

In general, the compounds of formula I and IA can be prepared using standard chemical reactions known in the art and are illustrated in Scheme A; wherein the terms K, P₄, P₃, P₂, R₁, R₂ and X have the same meanings as in formulas I and IA. Scheme A

In general, the acylated enols of formulas I and IA can be formed by reacting the peptide of formula 4 with a suitable symmetrical anhydride 2 or a suitable mixed anhydride 3 (wherein R₂' and R₂ are different but both are R₂ as indicated above) in the presence of an amine base such as the tertiary amines triethylamine and N-methylnorpholine or aromatic amines such as 4-dimethylaminopyridine, as well as picolines, collidines and pyridine. The reactants can be contacted with a suitable organic solvent such as acetonitrile, methylene chloride and the like. The reaction is usually carried out for a period of time ranging from about 30 minutes to about 48 hours at a temperature within the range of from about -40°C to about 85°C. In general, temperatures below 0ºC provide high ratios of IA to I and IA can be isolated in its pure form by chromatography or recrystallization. In Generally, reaction temperatures higher than 0 °C provide increasing ratios of I to IA and I can be isolated by chromatography and recrystallization.

Alternatively, the acylated enols of formulas I and IA can be formed by reacting the peptide of formula 4 with a suitable acid halide of formula R2-C(=O)X (X = F, Cl, Br, I) in the presence of a weakly basic amine such as picoline, collidine or pyridine.

Compounds of formula 4 are disclosed in European Patent Application Publication No. 0195212, inventor Michael Kolb et al., with a publication date of September 24, 1986, and in Peet, N. P. et al., J Med. Chem. 33 (1990), 394-407, both of which are hereby incorporated by reference as fully set forth.

Those compounds of formula 4 which are given here but not disclosed in European Patent Application Publication No. 0195212 or Peet, N. P. et al., J Med. Chem. 33 (1990), 394-407, can be prepared by the following synthetic procedures which are known and understood by those of ordinary skill in the art.

In general, all compounds of formula 4 can be prepared using standard chemical reactions known in the art and illustrated in Scheme B. Scheme B

Scheme B provides a general synthetic scheme for preparing the compounds of Formula 4.

Residues P2, P3 and K-P4 can be attached to the free amino group of the amino acid derivative of structure 5. Note that structure 5 represents the P1 moiety where the free carboxylic acid group has been substituted with an "X" moiety as indicated above. Residues P2, P3 and K-P4 can be attached to the unprotected, free amino compound (P1-X) by well-known peptide capping techniques. Furthermore, residues P1, P2, P3 and K-P4 can be joined together in any order so long as the final compound K-P4 is P3-P2-P1-X. For example, K-P4 can be attached to P3. to give K-P4-P3 which is attached to P2-P1-X, or K-P4 can be attached to P3-P2 which is then attached to an appropriate C-terminal protected residue P1 and the C-terminal protecting group is converted to X.

In general, peptides are extended by deprotecting the α-amine of the N-terminal residue and coupling the next appropriate N-protected amino acid through a peptide bond using the methods described. This deprotection and coupling procedure is repeated until the desired sequence is obtained. This coupling can be carried out with the constituent amino groups stepwise as illustrated in Scheme B, or by condensation of the fragments (two to several amino acids), or by a combination of both methods, or by solid phase peptide synthesis according to the method originally described by Marrifield, J. Am. Chem. Soc. 85 (1963), 2149-2154, the disclosure of which is hereby incorporated by reference. If a solid phase synthesis method is used, the C-terminal carboxylic acid is attached to an insoluble support (usually polystyrene). These insoluble supports contain a group that reacts with the carboxylic acid group to form a bond that is stable under the extension conditions but is easily cleaved later. Examples of these are: chloro- or bromomethyl resin, hydroxymethyl resin and aminomethyl resin. Many of these resins are commercially available with the desired C-terminal amino acid already incorporated.

In another embodiment, the compounds of the invention can be synthesized using an automated peptide synthesizer. In addition to the foregoing, peptide syntheses are described in Stewart and Young, "Solid Phase Peptide Synthesis," 2nd Edition, Pierce Chemical Co., Rockford, IL (1984); Gross, Meienhofer, Udenfriend, eds., "The Peptides: Analysis, Synthesis, Biology," vols. 1, 2, 3, 5 and 9, Academic Press, New York, 1980-1987; Bodanszky, "Peptide Chemistry: A Practical Textbook," Springer-Verlag, New York (1988); and Bodanszky et al., "The Practice of Peptide Synthesis," Springer-Verlag, New York (1984), the disclosures of which are hereby incorporated by reference.

The coupling between two amino acids, an amino acid and a peptide or two peptide fragments can be carried out using standard coupling methods such as the azide method, the mixed carbonic anhydride method (isobutyl chloroformate), the carbodiimide method (dicyclohexyl carbodiimide, diisopropyl carbodiimide or water-soluble carbodiimide), the active ester method (p-nitrophenyl ester, N-hydroxysuccinic acid imidoester), the Woodward reagent K method, the carbonyldiimidazole method, phosphorus reagents such as POP-Cl or the oxidation-reduction method. Some of these methods (especially the carbodiimide method) can be enhanced by adding 1-hydroxybenzotriazole. These coupling reactions can be carried out either in solution (liquid phase) or in solid phase.

The functional groups of the amino acids as essential components must generally be protected during the coupling reactions in order to avoid the formation of undesirable bonds. The protecting groups which can be used are listed in Greene, "Protective Groups in Organic Chemistry", John Wiley & Sons, New York (1981), and "The Peptides: Analysis, Synthesis, Biology" Vol. 3, Academic Press, New York (1981), and the disclosure of which is hereby incorporated by reference.

The α-carboxylic acid group of the C-terminal residue is usually, but not necessarily, protected by an ester which can be cleaved to yield the carboxylic acid.

Protecting groups which may be used include: 1) alkyl esters such as methyl and t-butyl esters, 2) aryl esters such as benzyl and substituted benzyl esters, or 3) esters which can be cleaved by treatment with weak base or weak reducing agents such as trichloroethyl or phenacyl esters.

The α-amino group of each amino acid to be coupled to the growing peptide chain must be protected. Any protecting group known in the art can be used. Examples of these include: 1) acyl types such as formyl, trifluoroacetyl, phthalyl and p-toluenesulfonyl group; 2) aromatic carbamate types such as benzyloxycarbonyl group (Cbz or Z) and substituted benzyloxycarbonyl groups, 1-(p-biphenyl)-1-methylethoxycarbonyl and 9-fluorenylmethyloxycarbonyl group (Fmoc); 3) aliphatic carbamate types such as tert-butyloxycarbonyl (Boc), ethoxycarbonyl, diisopropylmethoxycarbonyl and allyloxycarbonyl group; 4) cyclic alkyl carbamate types such as cyclopentyloxycarbonyl and adamantyloxycarbonyl group; 5) alkyl types such as triphenylmethyl and benzyl group; 6) trialkylsilane such as trimethylsilane; and 7) thiol-containing types such as phenylthiocarbonyl and dithiasuccinoyl group. The preferred α-amino protecting group is either Boc or Fmoc, preferably Boc. Many amino acid derivatives adequately protected for peptide synthesis are commercially available.

The α-amino protecting group of the newly added amino acid residue is cleaved prior to coupling of the next amino acid. If the Boc group is used, the reagent of choice is trifluoroacetic acid, neat or in dichloromethane, or HCl in dioxane or ethyl acetate. The resulting ammonium salt is then neutralized either prior to coupling or in situ with basic solutions such as aqueous buffers or tertiary amines in dichloromethane or dimethylformamide. If the Fmoc group is used, the reagents of choice are piperidine or substituted piperidine in dimethylformamide, but any secondary amine or basic solutions can be used. Removal of the protecting groups is carried out at a temperature between OCC and room temperature.

Each of the functionalities of the side chains bearing an amino acid must be protected during the preparation of the peptide using one of the groups described above. It will be understood by those skilled in the art that the selection and use of suitable protecting groups for these side chain functionalities depends on the amino acid and the presence of other protecting groups in the peptide. When selecting such protecting groups, it must be taken into account that they must not be removed during the deprotection and coupling of the α-amino group.

For example, if Boc is used as an α-amino protecting group, the following side chain protecting groups are suitable: p-toluenesulfonyl moieties (tosyl) can be used to protect the amino side chains of amino acids such as Lys and Arg, p-methylbenzyl, acetamidomethyl, benzyl (Bzl) or t-butylsulfonyl moieties can be used to protect the sulfide-containing side chains of amino acids such as cysteine, and benzyl (Bzl) ether can be used to protect the hydroxy-containing side chains of amino acids such as Ser or Thr.

When Fmoc is chosen for α-amino protection, tert-butyl-based protecting groups are usually acceptable. For example, Boc can be used for lysine, tert-butyl ether for serine and threonine, and tert-butyl ester for glutamic acid.

Once the peptide extension is complete, all protecting groups are removed. If solution phase synthesis is used, the protecting groups are removed in a manner dictated by the choice of protecting groups. These methods are well known to those skilled in the art.

When solid phase synthesis is used, the peptide is usually cleaved from the resin at the same time as the protecting groups are removed. When the Boc protection scheme is used in the synthesis, treatment with anhydrous HF containing additives such as dimethyl sulfide, anisole, thioanisole or p-cresol at 0ºC is the preferred method for cleavage of the peptide from the resin. Cleavage of the peptide can also be achieved by other acidic reagents such as trifluoromethanesulfonic acid/trifluoroacetic acid mixtures. When the Fmoc protection scheme is used, the N-terminal Fmoc group is the reagents described previously. Other protecting groups and the peptides are cleaved from the resin using a solution of trifluoroacetic acid and various additives such as anisole, etc.

Alternatively, the compounds of formula 4 can be prepared using standard chemical reactions known in the art and illustrated in Scheme C. Scheme C

Scheme C provides a general synthetic scheme in another embodiment for preparing the compounds of formula 4.

The residues P2, P3 and K-P4 can be attached to the free amino group of the amino alcohol derivative of structure 6 as described above in Scheme B to obtain the peptide alcohol of structure 7.

The alcohol functionality of the peptide alcohol of structure 7 is then oxidized by techniques and procedures known and understood by those of ordinary skill in the art, such as a Swern oxidation using oxalyl chloride and dimethyl sulfoxide to yield the compounds of formula 4.

The starting materials for use in Schemes B and C are readily available to one of ordinary skill in the art. For example, amino acids P2, P3 and KP4, where K is hydrogen, are commercially available. In addition, the amino protecting group K, where K is acetyl, succinyl, benzoyl, t-butyloxycarbonyl, carbobenzyloxy, dansyl, isovaleryl, methoxysucciny1, 1-adamantanesulfonyl, 1-adamantaneacety1, 2-carboxybenzoyl, 4-((chlorophenyl)sulfonylaminocarbonyl)phenylcarbonyl, 4-((4-bromophenyl)sulfonylaminocarbonyl)phenylcarbonyl and 4-(sulfonylaminocarbonyl)phenylcarbonyl, is described in European Patent Application Publication No. 363,284, published April 11, 1990, and U.S. Pat. No. 4,910,190, issued March 20, 1990, both of which are incorporated herein by reference as fully set forth. In addition, the radicals K, where K is -C(O)N(CH₃)₂ or a radical of the formulas:

or

where Z is N or CH, B is a radical of the formulas

or

R' is hydrogen or (C1-C4)alkyl, n is 0 or the integer 1 or 2, X is N or CH, in Angelastro, M. R. et al., J. Med. Chem. 37 (1994), 4538-4553, and in European Patent Application Publication No. 529,568, published March 3, 1993 and PCT International Publication No. WO 95/09838, published April 13, 1995, all three of which are hereby incorporated by reference as fully set forth. Synthetic methods for converting the K radicals into K-P4 substituents are known and understood by those of ordinary skill in the art.

Amino compounds as starting materials of formula 5 are readily available to one of ordinary skill in the art. For example, certain protected amino compounds of formula 5, where X is as indicated above, are described in European Patent Application Publication No. 195,212, published September 24, 1986. The publication is incorporated herein by reference as fully set forth.

In addition, other starting materials for use in Schemes B and C can be prepared by the following synthetic procedures, which are known and understood by those of ordinary skill in the art.

The production process of substituted amino acids KP₄, where K

wherein B is a -C(=O)- group and is outlined in Scheme D, wherein P₄ and Z are as defined above or the functional equivalents of these groups. Scheme D

Specifically, the amino acids KP₄, where K

wherein B is a -C(=O) group, by coupling the amino acid KP₄, wherein K is a hydrogen atom, with an acid chloride of structure 8 in the presence of one to four molar equivalents of a suitable amine which can act as a hydrogen halide acceptor. Suitable amines for use as hydrogen halide acceptors are tertiary organic amines such as tri(lower alkyl)amines, e.g. triethylamine, or aromatic amines such as picolines, colloidines and pyridine. If pyridines, picolines or coilidines are used, they can be used in large excess and therefore also act as reaction solvents. N-methylnorpholine ("NMM") is particularly suitable for the reaction. The coupling reaction can be carried out by adding an excess, such as a 1-5, preferably about 4-fold, molar excess of the amine and then the acid chloride of structure 8 to a solution of the amino acid KP4, where K is hydrogen. The solvent can be any suitable solvent, e.g., petroleum ether, a chlorinated hydrocarbon such as carbon tetrachloride, ethylene chloride, methylene chloride or chloroform, a chlorinated aromatic such as 1,2,4-trichlorobenzene or o-dichlorobenzene, carbon disulfide, an ethereal solvent such as diethyl ether, tetrahydrofuran or 1,4-dioxane, or an aromatic solvent such as benzene, toluene or xylene. The preferred solvent for this coupling reaction is methylene chloride. The reaction proceeds for about 15 minutes to about 6 hours, depending on the reactants, solvent, concentrations and other factors, again at a temperature which may be from about 0ºC to about 60ºC and is conveniently about room temperature, ie 25ºC. The N-protected amino acids KP₄, where K

where B is a -C(=O) group, can be isolated from the reaction mixture by any suitable technique, such as by chromatography on silica gel.

The substituted amino acids KP₄, where K

where B is different from a -C(=O) group, can be prepared analogously only by substituting the appropriate intermediate

where B is other than a -C(=O) group and A is Cl or OH (the corresponding acid, acid chloride or sulfonyl chloride) with the compound of structure 8 in Scheme D.

The acid chloride of structure 8 and the appropriate intermediate of formula

where 13 is other than a -C(=O) group and A is Cl or OH, are commercially available or can be readily prepared by techniques and procedures known and understood by those of ordinary skill in the art.

For example, the appropriate intermediate of the formula

as outlined in Scheme E, with all substituents as indicated above. Scheme E

Scheme E provides a general synthetic procedure for preparing the appropriate intermediate of formula

ready.

In step a, the carboxylic acid functionality of the appropriate 2,5-pyridinedicarboxylic acid 2-methyl ester 10 (Nippon Kagaku Zasshi, 88 (1967), 563) is converted to its acid chloride using techniques and procedures known and understood by those of ordinary skill in the art, such as with thionyl chloride, to yield the corresponding 6-carbomethoxynicotinoyl chloride 11.

In step b, the acid chloride is amidated with morpholine 12 by techniques and procedures known and understood by those of ordinary skill in the art to obtain the corresponding 5-(morpholine-4-carbonyl)-2-pyridinecarboxylic acid methyl ester 13.

In step c, the methyl ester functionality of 13 is hydrolyzed by techniques and methods known and understood by those of ordinary skill in the art, e.g., with lithium hydroxide in methanol, to yield 5-(morpholine-4-carbonyl)-2-pyridinecarboxylic acid 14.

In addition, the appropriate intermediate of the formula

as outlined in Scheme F, with all substituents as indicated above. Scheme F Scheme F

Scheme F provides a general synthetic procedure for preparing the appropriate intermediate of formula

ready.

In step a, the free carboxylic acid functionality of 2,5-pyridinecarboxylic acid 2-methyl ester 10 (Nippon Kagaku Zasshi, 88 (1967), 563) is converted to its t-butyl ester using techniques and procedures known and understood by those of ordinary skill in the art, such as by the t-butyl alcohol adduct of dicyclohexylcarbodiimide (Synthesis (1997), 570), to obtain the corresponding 2,5-pyridinedicarboxylic acid 2-methyl ester 5-t-butyl ester 15.

For example, 2,5-pyridinedicarboxylic acid 2-methyl ester 10 is combined with a molar excess of the t-butyl alcohol adduct of dicyclohexylcarbodiimide in a suitable organic solvent such as methylene chloride. The reaction is usually in a temperature range of 0ºC to room temperature and for a time period in the range of 2-24 hours. The 2,5-pyridinedicarboxylic acid 2-methyl ester 5-t-butyl ester 15 is isolated from the reaction mixture by standard extraction procedures as known in the art and can be purified by crystallization.

In step b, the methyl ester functionality of 15 is amidated with morpholine 12 to give the corresponding 6-(morpholine-4-carbonyl)nicotinic acid t-butyl ester 16.

For example, 2,5-pyridinedicarboxylic acid 2-methyl ester 5-t-butyl ester 15 is contacted with a molar excess of morpholine in a suitable organic solvent such as tetrahydrofuran. The reaction is usually carried out at a temperature range from room temperature to reflux and for a time period in the range from 5 hours to 3 days. The 6-(morpholine-4-carbonyl)nicotinic acid t-butyl ester 16 is isolated from the reaction mixture by standard extraction procedures as known in the art and can be purified by crystallization.

In step c, the t-butyl ester functionality of 16 is hydrolyzed, e.g. with HCl in nitromethane, to afford the corresponding 6-(morpholine-4-carbonyl)nicotinic acid 17.

In general, the compounds of formula 4 or 5 can be prepared using standard chemical reactions analogous to those known in the art. Compounds of formula 5 or 6, wherein X is -CO₂R₃, i.e. wherein the compounds are α-ketoesters, can be prepared as described by Angelastro, M. R. et al., J. Med. Chem., 33 (1990), 11, Peet, N. P. et al., J. Med. Chem., 33(1990), 394, Mehdi, S. et al., Biochem. Biophys. Res. Comm., 166 (1990), 595, and European Patent Application OPI No. 0195212, inventors Michael Kolb et al., with a publication date of September 24, 1985, and all four references are hereby incorporated by reference as fully set forth.

The compounds of formula 4 or 5, wherein X is -CONHR₃, i.e. wherein the compounds are α-ketoamides, can be prepared as described in PCT International Publication No. WO 95/C19838, published April 13, 1995, hereby incorporated by reference as fully set forth. The compounds of formula 4 or 5 wherein X is -CONHR₃ may also be prepared by the procedure described in Scheme G. All substituents are as defined above unless otherwise indicated. The reagents and starting materials are readily available to one of ordinary skill in the art. Scheme G

The required α-ketoester starting material 18 can be prepared as described by Angelastro, MR et al., J. Med. Chem., 33 (1990), 11, Peet, NP et al., J. Med. Chem., 33 (1990), 394, and European Patent Application OPI No. 0195212, inventors Michael Kolb et al., with a Publication date of September 24, 1986. The term "Pg" refers to an appropriate protecting group as described in more detail above.

In Scheme G, step a, the α-keto ester 18 is selectively hydrolyzed to an α-keto acid by treatment with an appropriate base.

For example, the appropriately substituted α-keto ester 18 is dissolved in a suitable solvent mixture, such as methanol:water (50:50), and treated with one equivalent of a suitable base, such as lithium hydroxide. The reaction is stirred at a temperature of about 0°C to 30°C for about 1 hour to 10 hours. The α-keto acid 19 is then isolated by extraction techniques known in the art. For example, the reaction mixture is diluted with a suitable organic solvent, such as ethyl acetate, and an equal volume of water. The layers are separated. The aqueous layer is acidified with dilute hydrochloric acid and extracted with a suitable organic solvent, such as ethyl acetate. The combined organic extracts are dried over anhydrous magnesium sulfate, filtered, and concentrated under vacuum to obtain the α-keto acid 19.

In Scheme G, step b, the α-keto acid 19 is coupled with a primary amine 20 under conditions known in the art to afford the desired α-ketoamide 21.

For example, the appropriately substituted α-keto acid 19 is dissolved in a suitable organic solvent such as methylene chloride. The solution is then treated with one equivalent of 1-hydroxybenzotriazole, one equivalent of diisopropylethylamine and an excess of a primary amine 20. One equivalent of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride is added and the reaction mixture is stirred at a temperature of about 0°C to 25°C for about 2 to 10 hours. The product is then isolated by techniques known in the art. For example, the reaction mixture is diluted with ethyl acetate, washed with cold 0.5 N hydrochloric acid, saturated sodium bicarbonate solution, dried over anhydrous magnesium sulfate, filtered, and concentrated in vacuo to provide the α-ketoamide 21.

In Scheme G, step c, the protecting groups of the α-ketoamide 21 are removed under conditions known in the art as described in T. H. Green "Protective Groups in organic Synthesis", John Wiley & Sons, 1981, Chapter 7 to provide the unprotected α-ketoamide 22. For example, if "Pg" is a t-butyloxycarbonyl group (Boc), the α-ketoamide 21 is dissolved in a suitable solvent such as ethyl acetate, treated with an excess of hydrogen chloride (gas) and stirred at about 0°C to 30°C for about 30 minutes to 4 hours. The solvent is then removed in vacuo to provide the unprotected α-ketoamide 22 as an HCl salt.

The unprotected α-ketoamide 22 is then subjected to the reaction conditions described in Scheme B to provide the compounds of formula 23.

The following examples represent typical syntheses. These examples are intended to be illustrative only and are not intended to limit the scope of the present invention in any way. As used herein, the following terms have the meanings indicated: "g" refers to grams, "mmol" refers to millimoles, "ml" refers to milliliters, "bp" refers to boiling point, "ºC" refers to degrees Celsius, "mmHg" refers to millimeters of mercury, "ul" refers to microliters, "ug" refers to micrograms, and "uM" refers to micromolar. Example 1 Preparation of L-prolinamide, N-(4-methoxy-1,4-dioxobutyl)-L-alanyl-L-alanyl-N-[2-(acetyloxyl-3-methoxy-1-(1-methylethyl)-3-oxo-1-propenyl]-, (E)- and (Z)-mixture (SEQ. ID NO: 6)

Scheme A: To pyridine (1.25 mL) stirred under N2 and cooled to -20 °C is added MeOSuc-Ala-Ala-Pro-Val-CO2-CH3 (156 mg, 0.30 mmol) (Peet, N. P. et al., J. Med. Chem. 33 (1990), 394 or Angelastro, M. R. et al., J. Med. Chem. 33 (1990), 11), and a few minutes later acetic anhydride (0.29 mL, 3.0 mmol). The reaction mixture is warmed to room temperature and stirred for 20 hours. The reaction mixture is diluted with CH2Cl2 (30 mL) and washed with 0.3 N HCl (2 x 20 mL) followed by brine (15 mL). Drying (MgSO4), filtration and concentration afforded a crude product. Flash chromatography (4 x 14 cm silica gel column) using a gradient (25 to 50%) of acetone in EtOAc as eluent afforded the title compound (88 mg, 53%, E:Z isomer ratio 9:1) as a colorless oil.

1 H-NMR (CDCl3 , 400 MHz) ? 10.16 (br s, 0.9H, CONHC = C of the E isomer), 8.23 (br s, 0.1H, CONHC=C of the Z isomer), 7.49 (br d, 0.1H , NH of the Z-isomer), 7.17 (br d, 0.9H, NH of the E- isomer), 6.50 (br d, 0.1H, NH of the Z-isomer), 6.43 (br d , 0.9H, NH of E-isomer), 4.81- 4.72 (m, 1.1H, CH of Ala for E-isomer and CH of Ala for Z-isomer and CH of Pro for Z-isomer) , 4.64 (t 0.1H, CH of Ala for Z-isomer), 4.58-4.48 (m, 1.8H, CH of Ala for E-isomer and CH of Pro for E-isomer), 3.76-3.63 (m, 2.7H, CH₂N of both isomers and 2 · OCH₃ of Z -isomer and CH of the Z-isomer), 3.42 (septet, 0.9H, CH of the E-isomer), 3.68 (s, 2.7H, OCH₃ of the E-isomer), 3.66 (s , 2.7H, OCH3 of the E isomer), 2.68-2.60 and 2.53-2.45 (pr m, 4H, COCH₂CH₂CO of both isomers), 2.22-1.95 (m, 4H, CH₂CH₂ of both isomers), 2.20 (s, 3H, CH₃CO of both isomers), 1, 37 (d, 2.7H, CH3 of Ala for E-isomer), 1.32 (d, 2.7H, CH3 of Ala for E-isomer), 1.27 (d, 0, 3H, CH3; of Ala for Z-isomer), 1.25 (d, 0.3H, CH₃ of Ala for Z-isomer), 1.14 and 1.13 (pr d, 5.4H, 2 x CH₃ of Val for E-isomer), 1.05 and 1.02 (pr d, 0.6H, 2 x CH₃ of Val for Z-isomer).

MS (CI, CH4) m/z (rel. intensity) 555 (MH+, 62), 354 (38), 299 (100).

HRMS C₂₅H₃₉N₄O₁₀ (MH⁺) calcd 555.2666, obs 555.2669. Example 2 Preparation of L-prolinamide, N-[4-[[[(4-chlorophenyl)sulfonyl]amino]carbonyl]benzoyl]-Lvalyl-N-[2-(acetyloxy)-3-methoxy-1-(1-methylethyl)-3-oxo-1-propenyl]-, (E)-

Scheme A: N-(4-((4-Chlorophenyl)sulfonylaminocarbonyl)phenylcarbonyl-Val-Pro- Val-CO₂CH₃; (135 mg, 0.20 mmol) (Mehdi, S.- et al., Biochem. Biophys. Res. Comm. 166 (1990), 595) is treated with acetic anhydride (0.19 mL, 2.0 mmol) in pyridine (1.0 mL), followed by preparative TLC (developed using EtOAc) in a manner analogous to that described in Example 1 to afford the title compound (23 mg, 16%) as a white foam.

1 H-NMR (CDCl3 , 300 MHz) ? 10.40 (br s, 1H, CONHC=C), 8.11-8.05 (m, 2H, aryl), 7.78-7.71 (m, 2H, aryl), 7.71-7, 63 (m, 2H, aryl), 7.55-7.49 (m, 2H, aryl), 7.24 (br d, 1H, NH of Val, 4.98 (dd, 1H, CH), 4.46 (dd, 1H, CH), 4.02-3.89 and 3.89-3.76 (pr m, 2H, CH₂N), 3, 68 (s, 3H, (OCH₃), 3.47 (septet 1H, CHC=C), 2.39-1.96 (m, 5H, CH₂CH₂ and CH), 2.21 (s, 3H , CH₃CO), 1.17 and 1.16 and 1.07 and 0.94 (four d, 12H, 4 · CH₃).

MS (CI, CH4) m/z (rel. intensity) 749 (1), 747 (M+C2H5, 2), 721 (4), 719 (M+H, 10), 299 (100).

HRMS C₃₃H₄�0ClN₄O₁₀S (MH⁺) calcd 719.2154, obs 719.2146. Example 3 Preparation of L-prolinamide, N-[4-[[[(4-chlorophenyl)sulfonyl]aminolcarbonyl]benzoyl]-Lvalyl-N-[2-(acetyloxy)-3-amino-1-(1-methylethyl)-3-oxo-1-propenyl]-, (E)-

a) Preparation of N-(4-((4-chlorophenyl)sulfonylaminocarbonyl)phenylcarbonyl-Val- Pro -Val-COOH

Scheme G, step a: N-(4-((4-chlorophenyl)sulfonylaminocarbonyl)phenylcarbonyl- Val-Pro-Val-CO₂CH₃ (677 mg, 1.0 mmol) was dissolved in a solvent mixture of THF:methanol:water (1:1:1) (30 mL) and treated with 1.0 N aqueous lithium hydroxide solution (2.2 mL, 2.2 mmol). The reaction mixture was stirred at 0 °C for 3 h, then the reaction mixture was diluted with ethyl acetate (100 mL) followed by an equal volume of water. After separating the layers, 1 N HCl (3 mL) was slowly added to the aqueous layer and extracted with ethyl acetate (3 x 25 mL). The combined organic layers are dried over anhydrous magnesium sulfate, filtered and concentrated in vacuo to obtain the title compound

b) Preparation of N-(4-((4-chlorophenyl)sulfonylaminocarbonyl)phenylcarbonyl-Val- Pro -Val-CONH2;

Scheme G, step b: The product of Example 3(a) is dissolved in methylene chloride (20 mL). Then, 1-hydroxybenzorazole (68 mg, 0.5 mmol), diisopropylethylamine (0.17 mL, 1.0 mmol) are added to the reaction mixture and three equivalents of anhydrous ammonia (26 mg, 1.5 mmol) are bubbled through. Then, 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (96 mg, 0.5 mmol) is added to the reaction mixture and stirred at a temperature of 0 °C for 4 hours. The reaction mixture is diluted with methylene chloride (10 mL), washed with cold 0.5 N HCl (15 mL) and saturated aqueous sodium bicarbonate solution (15 mL). Then it is dried over anhydrous magnesium sulfate, filtered and concentrated in vacuo to obtain the title compound.

c) Preparation of L-prolinamide, N-[4-[[[(4-chlorophenyl)sulfonyl]amino]carbonyl] benzoyl]-L-valyl-N-[2-(acetyloxy)-3-amino-1-( 1-methylethyl)-3-oxo-1-propenyl]-, (E)-

Scheme A: The product of Example 3(b) (141 mg, 0.20 mmol) is treated with acetic anhydride (0.19 mL, 2.0 mmol) in pyridine (1.0 mL) followed by preparative TLC in a manner analogous to that described in Example 1 to afford the title compound. Example 4 Production of L-prolinamide, N-[4-[[[(4-chlorophenyl)sulfonyl]amino]carbonyl]benzoyl]-Lvalyl- N-[1-(1-methylethyl)-3-oxo-2-(acetyloxy)-3-(phenylamino)-1-propenyl]-, (E)-

a) Preparation of N-(4-((4-chlorophenyl)sulfonylaminocarbonyl)phenylcarbonyl-Val- Pro -Val-CONHC₆H₅

Scheme G, step b: Aniline (46 μl, 0.5 mmol) is coupled to the product of Example 3(a) in a manner analogous to the procedure of Example 3(b) to give the title product.

b) Preparation of L-prolinamide, N-[4-[[[(4-chlorophenyl)sulfonyl]amino]carbonyl]-benzoyl]-L-valyl-N-[1-(1-methylethyl)-3-oxo- 2-(acetyloxy)-3-(phenylamino)-1-propenyl]-, (E)-

Scheme A: The product of Example 4(a) (156 mg, 0.20 mmol) is treated with acetic anhydride (0.19 mL, 2.0 mmol) in pyridine (1.0 mL) followed by preparative TLC in a manner analogous to that described in Example 1 to afford the title compound. Example 5 Preparation of L-prolinamide, N-acetyl-L-valyl-N-[-2-(acetyloxy)-3-ethoxy-1-(1-methylethyl)-3-oxo-1-propenyl]-, (E)-

a) Preparation of ethyl 3-amino-2-hydroxy-4-methylpentanoate hydrochloride

Treat 3-Amino-2-hydroxy-4-methylpentanoic acid (0.89 g, 6.05 mmol (Peet, N. P. et al., J. Med. Chem. 33 (1990), 394) in CH₃CH₂OH (25 mL and then 20 mL) with gaseous HCl. Concentrate and dry over KOH pads to give the title compound.

b) Preparation of 3-[(N-acetyl-L-valyl-L-prolyl)aminol-2-hydroxy-4-methylpentanoic acid ethyl ester

A solution of NMM (0.22 mL, 2.00 mmol) and Ac-Val-Pro-OH (531 mg, 2.00 mmol) in CH3CN (20 mL) is cooled to -20 °C and i-BuOCOCl (26 mL, 2.00 mmol) is added. After 10 min, a solution of the product of Example 5(a) (423 mg, 2.00 mmol) and N-methylmorpholine (0.22 mL, 2.0 mmol) in CHCl3 (3 mL) is added and the reaction mixture is slowly warmed to room temperature. After 2.5 h, the reaction mixture is diluted with methylene chloride, washed with 0.5 N HCl (2 x 15 mL) and saturated aqueous NaHCO3 (2 x 15 mL). Dry (MgSO4), filter and concentrate to give a crude product. Flash chromatography to give the title compound.

c) Preparation of N-acetyl-L-valyl-N-[3-ethyoxy-1(1-methylethyl)-2,3-dioxopropyl]- L-prolinamide

To a stirred solution of oxalyl chloride (0.12 mL, 1.43 mmol) in CH2Cl2 (1.5 mL) cooled to -60°C is added DMSO (0.20 mL, 2.86 mmol) in CH2Cl2 (0.5 mL). After 5 min, the product of Example 5(b) (298 mg, 0.72 mmol) in CH2Cl2 (1.5 mL) is added. Stirring is continued for 15 min at -60°C. After adding Et3N (0.50 mL, 3.58 mmol), the reaction mixture is warmed to room temperature and directly added to a column for flash chromatography. Fractions containing product are combined and concentrated to give the title compound.

d) Preparation of L-prolinamide, N-acetyl-L-valyl-N-[2-(acetyloxy)-3-ethoxy-1-(1-methylethyl)-3-oxo-1-propenyl]-, (E)-

Scheme A: The product of Example 5(c) (83 mg, 0.20 mmol) is treated with acetic anhydride (0.19 mL, 2.0 mmol) in pyridine (1.0 mL) followed by preparative TLC in a manner analogous to that described in Example 1 to afford the title compound. Example 6 Preparation of L-prolinamide, N-acetyl-L-prolyl-L-valyl-N-[2-(acetyloxy)-3-methoxy-1-(1-methylethyl)-3-oxo-1-propenyl]-, (E)-(SEQ. ID. NO. 7)

Scheme A: N-Acetyl-L-prolyl-L-alanyl-N-(3-methoxy-1-methyl-2,3-dioxopropyl)-Lprolinamide (88 mg, 0.20 mmol (Peet, NP et al., J. Med. Chem. 33 (1990), 394) is treated with acetic anhydride (0.19 mL, 2.0 mmol) in pyridine (1.0 mL), followed by preparative TLC, in a manner analogous to that described in Example 1, to obtain the title compound. Example 7 Preparation of L-prolinamide, N-[4-(4-morpholinylcarbonyl)benzoyl]-L-valyl-N-[2-(acetyloxy)-3-ethoxy-1-(1-methylethyl)-3-oxo-1-propenyl]-, (E)-

a) Preparation of 3-[(N-Boc-L-valyl-L-prolylamino-2-hydroxy-4-methylpentanoic acid ethyl ester

Boc-Val-Pro-OH (630 mg, 2.00 mmol) is coupled to the product of Example 5(a) (423 mg, 2.00 mmol) in a manner analogous to that described in Example 5(b) to afford the title compound.

b) Preparation of N-Boc-L-valyl-N-[3-ethyloxy-1-(1-methylethyl)-2,3-dioxopropyl]-Lprolinamide

The product of Example 7(a) (338 mg, 0.72 mmol) is oxidized in a manner analogous to that described in Example 5(c) to afford the title compound.

c) Preparation of N-L-valyl-N-[3-ethyloxy-1-(1-methylethyl)-2,3-dioxopropyl]-L-prolinamide hydrochloride salt

HCl gas is bubbled through a cold (0 °C) solution of the product of Example 7(b) (0.94 g, 2.00 mmol) in EtOAc (50 mL) for 5 min. The reaction mixture is stirred at 0 °C for 1.5 h and the solvent is removed in vacuo to provide the title compound, which is used without further purification.

d) Preparation of N-[4-(4-morpholinylcarbonyl)benzoyl]-L-valyl-N-[3-ethoxy-1-(1-methylethyl)-2,3-dioxopropyl]-L-prolinamide

To a stirred suspension of 4-(4-morpholinylcarbonyl)benzoic acid (235 mg, 1.0 mmol) (Angelastro, M. R. et al., J. Med. Chem. 37 (1994), 4538) and benzyltriethylammonium chloride (2 mg) in 1,2-dichloroethane (5 mL) is added thionyl chloride (88 µL, 1.2 nimol) and heated to reflux. After 19 hours, the solution is cooled to room temperature and concentrated to give the acid chloride as a light orange liquid, which is used without further purification. In a separate flask, a stirred solution of the product of Example 7(c) (406 mg, 3.0 mmol) in CH2Cl2 (10 mL) is cooled to -20 °C. NMM (0.33 mL, 3.0 mmol) is added, immediately followed by the addition of a solution of the acid chloride in CH2Cl2 (5 mL) at a rate to maintain the internal reaction temperature at -13°C or less. After the addition is complete, the reaction mixture is warmed to room temperature. After an additional 2 h, the reaction mixture is diluted with CH2Cl2 (20 mL) and washed with 0.5 N HCl (2 x 15 mL), saturated NaHCO3 (2 x 15 mL), and brine (15 mL). Dry and concentrate to give the crude title compound. Flash chromatography to give the title compound.

e) Preparation of L-prolinamide, N-[4-(4-morpholinylcarbonyl)benzoyl]-L-valyl-N-[2-(acetyloxy)-3-ethoxy-1-(1-methylethyl)-3-oxo- 1-propenyl]-, (E)-

Scheme A: The product of Example 7(d) (117 mg, 0.20 mmol) is treated with acetic anhydride (0.19 mL, 2.0 mmol) in pyridine (1.0 mL) followed by preparative TLC in a manner analogous to that described in Example 1 to afford the title compound. Example 8 Preparation of L-prolinamide, N-(4-morpholinylcarbonyl)-L-valyl-N-[2-(acetyloxy)-3- ethoxy-1-(1-methylethyl)-3-oxo-1-propenyl]-, (E)-

a) Preparation of N-(4-morpholinylcarbonyl)-L-valyl-N-[3-ethoxy-1-(1-methylethyl)- 2,3-dioxopropyl]-L-propynamide

To a solution of the product of Example 7(c) (406 mg, 1.0 mmol) in CH2Cl2 (20 mL) is added 4-morpholinecarbonyl chloride (0.47 mL, 4.0 mmol) and NMM (0.55 mL, 5.0 mmol). The mixture is stirred for 2.5 hours, the solvent is evaporated and the residue is purified by flash chromatography to provide the title compound.

b) Preparation of L-prolinamide, N-[4-(4-morpholinylcarbonyl)benzoyl]-L-valyl-N-[2-(acetyloxy)-3-ethoxy-1-(1-methylethyl)-3-oxo- 1-propenyl]-, (E)-

Scheme A: The product of Example 8(a) (97 mg, 0.20 mmol) is treated with acetic anhydride (0.19 mL, 2.0 mmol) in pyridine (1.0 mL) followed by preparative TLC in a manner analogous to that described in Example 1 to afford the title compound. Example 9 Preparation of L-prolinamide, N-(2-furoyl)-L-valyl-N-[2-(acetyloxy)-3-ethoxy-1-(1-methylethyl)-3-oxo-1-propenyl]-, (E)-

a) Preparation of N-(2-furoyl)-L-valyl-N-[3-ethyloxy-1-(1-methylethyl)-2,3-dioxopropyliprolinamide

2-Furoyl chloride (0.10 mL, 1.0 mmol) is coupled with the product of Example 7(c) (406 mg, 1.0 mmol in the presence of NMM (0.33 mL, 3.0 mmol) in a manner analogous to that described in Example 8(a) to provide the title compound.

b) Preparation of L-prolinamide, N-(2-furoyl)-L-valyl-N-[2-(acetyloxy)-3-ethoxy-1-(1-methylethyl)-3-oxo-1-propenyl]-, (E)-

Scheme A: The product of Example 9(a) (93 mg, 0.20 mmol) is treated with acetic anhydride (0.19 mL, 2.0 mmol) in pyridine (1.0 mL) followed by preparative TLC in a manner analogous to that described in Example 1 to afford the title compound. Example 10 Preparation of L-prolinamide, N-[(tetrahydro-2H-pyran-4-yl)carbonyl]-L-valyl-N-[2-(acetyloxy)-3-ethoxy-1-(1-methylethyl)-3-oxo-1-propenyl]-, (E)-

a) Preparation of N-[(Tetrahydro-2H-pyran-4-yl)carbonyl]-L-valyl-L-valyl-N-[3-ethyloxy-1-(1-methylethyl)-2,3-dioxopropyl] -L-prolinamide

To a mixture of tetrahydro-2H-pyran-4-carboxylic acid (130 mg, 1.0 mmol) (Angelastro, M. R. et al., J. Med. Chem. 37 (1994), 4538) and DMF (0.1 mL) in CH2Cl2 is added oxalyl chloride (87 µL, 1.0 mmol). The mixture is stirred at room temperature for 0.5 h, followed by the addition of NMM (0.33 mL, 3.0 mmol) and the product of Example 7(c) (406 mg, 1.0 mmol). The mixture is stirred for 2.5 h, poured into H2O and extracted with methylene chloride. The extracts are combined, dried (MgSO4) and concentrated. The residue is purified by flash chromatography to give the title compound.

b) Preparation of L-prolinamide, N-[(tetrahydro)-2H-pyran-4-yl)cabonyl]-L-valyl-N-[2-(acetyloxy)-3-ethoxy-1-(1-methylethyl) -3-oxo-1-propenyl]-, (E)-

Scheme A: The product of Example 10(a) (96 mg, 0.20 mmol) is treated with acetic anhydride (0.19 mL, 2.0 mmol) in pyridine (1.0 mL) followed by preparative TLC in a manner analogous to that described in Example 1 to afford the title compound. Example 11 Preparation of L-prolinamide, N-[3-(4-morpholinyl)-1,3-dioxopropyl]-L-valyl-N-[2-(acetyloxy)-3-ethoxy-1-(1-methylethyl)-3-oxo-1-propenyl]-, (E)-

a) Preparation of N-[3-(4-morpholinyl)-1,3-dioxopropyl]-L-valyl-N-[3-ethyloxy-1-(1- methylethyl-2,3-dioxopropyl]-L-prolinamide

2-(4-Morpholinylcarbonyl)ethanoic acid (173 mg, 1.0 mmol) (Angelastro, M. R. et al., J. Med. Chem. 37 (1994), 4538) is activated with oxalyl chloride (87 μL, 1.0 mmol) and coupled with the product of Example 7(c) (406 mg, 1.0 mmol) in the presence of NMM (0.33 mL; 3.0 mmol) in a manner analogous to that described in Example 10(a). Purification by flash chromatography provides the title compound.

b) Preparation of L-prolinamide, N-[3-(4-morpholinyl)-1,3-dioxopropyl]-L-valyl-N-[2-(acetyloxy)-3-ethoxy-1-(1-methylethyl) -3-oxo-1-propenyl]-, (E)-

Scheme A: The product of Example 11(a) (105 mg, 0.20 mmol) is treated with acetic anhydride (0.19 mL, 2.0 mmol) in pyridine (1.0 mL) followed by preparative TLC in a manner analogous to that described in Example 1 to afford the title compound. Example 12 Preparation of L-prolinamide, N-(3-pyridyl)propanoyl]-L-valyl-N-[2-(acetyloxy)-3- ethoxy-1-(1-methylethyl)-3-oxo-1-propenyl]-, (E)-

a) Preparation of N-[3-(3-pyridyl)propanoyl]-L-valyl-N-[3-ethyloxy-1-(1-methylethyl)-2,3-dioxopropyl-L-prolinamide

To a suspension of 3-(3-pyridyl)propionic acid (302 mg, 2.0 mmol) (Walker, F. A. et al., J. Am. Chem. Soc. 102 (1980), 5530) in CH2Cl2 (30 mL) is added Et3N (0.84 mL, 6.0 mmol) and the resulting solution is cooled to -22 °C. IBCF (0.26 mL, 2.0 mmol) is added and the reaction mixture is stirred for 20 min. Additional Et3N (0.28 mL, 2.0 mmol) is added, followed by the addition of the product of Example 7(c) (812 mg, 2.0 mmol) in one portion. After stirring at -22ºC for 4 hours, the reaction mixture is concentrated and purified by flash chromatography to afford the title compound.

b) Preparation of L-prolinamide, N-[3-(3-pyridyl)propanoyl]-L-valyl-N-[2-(acetyloxy)-3-ethoxy-1-(1-methylethyl)-3-oxo- 1-propenyl]-, (E)-

Scheme A: The product of Example 12(a) (101 mg, 0.20 mmol) is treated with acetic anhydride (0.19 mL, 2.0 mmol) in pyridine (1.0 mL) followed by preparative TLC in a manner analogous to that described in Example 1 to afford the title compound. Example 13 Preparation of glycinamide, N-(4-methoxy-1,4-dioxobutyl)-L-valyl-N-[2-(acetyloxy)-3- methoxy-1-(1-methylethyl)-3-oxo-1-propenyl-N²-(2,3-dihydro-1H-inden-2-yl]-, (E)-

a) Preparation of CBZ-Val-N-(2,3-dihydro-1H-inden-2-yl)Gly-Val(OH)-CO₂CH₃

N-(Carbobenzyloxy)-L-valyl-N-(2,3-dihydro-1H-inden-2-yl)glycine (425 mg, 1.0 mmol) (Skiles, J. W. et al., J. Med. Chem. 35 (1992), 641) is coupled to methyl 3-amino-2-hydroxy-4-methylpentanoate (161 mg, 1.0 mmol) (Peet, N. P. et al., J. Med. Chem. 33. (1990), 394) in a manner analogous to the procedure of Example 5(b) to provide the title compound.

b) Preparation of CBZ-Val-N-(2,3-dihydro-1H-inden-2-yl)Gly-Val-CO₂CH₃

The title compound is prepared from the product of Example 13(a) (284 mg, 0.5 mmol) by following the oxidation procedure described in Example 5(c).

c) Preparation of glycinamide, N-(4-methoxy-1,4-dioxobutyl)-L-valyl-N-[2- (acetyloxy)-3-methoxy-1-(1-methylethyl)-3-oxo-1-propenyl-N²-(2,3-dihydro-1H-inden-2-yl]-, (E)-

Scheme A: The product of Example 13(b) (113 mg, 0.20 mmol) is treated with acetic anhydride (0.19 mL, 2.0 mmol) in pyridine (1.0 mL) followed by preparative TLC in a manner analogous to that described in Example 1 to afford the title compound. Example 14 Preparation of glycinamide, N-(4-methoxy-1,4-dioxobutyl)-L-valyl-N-[2-(acetyloxy)-3- methoxy-1-(1-methylethyl)-3-oxo-1-propenyl-N²-methyl]-, (E)-

a) Preparation of CBZ-Val-N-(methyl)Gly-Val(OH)-CO₂CH₃

N-(Carbobenzyloxy)-L-valyl-N-(methyl)glycine (322 mg, 1.0 mmol) (Skiles, J. W. et al., J. Med. Chem. 35 (1992), 641) is coupled to methyl 3-amino-2-hydroxy-4-methylpentanoate (161 mg, 1.0 mmol) (Peet, N. P. et al., J. Med. Chem. 33 (1990), 394) in a manner analogous to the procedure of Example 5(b) to provide the title compound.

b) Preparation of N-(carbobenzyloxy)-L-valyl-N-methyl-N-[3-methoxy-1-(1-methylethyl)-2,3-dioxopropyl]glycinamide

The title compound is prepared from the product of Example 14(a) (233 mg, 0.5 mmol) by following the oxidation procedure described in Example 5(c).

c) Preparation of glycinamide, N-(4-methoxy-1,4-dioxobutyl)-L-valyl-N-[2-(acetyloxy)-3-methoxy-1-(1-methylethyl)-3-oxo-1 -propenyl-N²-methyl]-, (E)-

Scheme A: The product of Example 14(b) (93 mg, 0.20 mmol) is treated with acetic anhydride (0.19 mL, 2.0 mmol) in pyridine (1.0 mL) followed by preparative TLC in a manner analogous to that described in Example 1 to afford the title compound. Example 15 Preparation of L-prolinamide, N-(4-methoxy-1,4-dioxobutyl)-L-alanyl-L-alanyl-N-[2-(acetyloxy)-3-methoxy-1-(1-methylethyl)-3-oxo-1-propenyl]-, (Z)- (SEQ. ID NO: 8)

Scheme A: To a stirred solution of pyridine (1.25 mL) and MeOSuc-Ala-Ala-Pro-Val-CO2CH3 (156 mg, 0.30 mmol) heated to reflux under N2 atmosphere was added dropwise acetic anhydride (0.29 mL, 3.0 mmol). After 30 min at reflux, the reaction mixture was cooled, diluted with CH2Cl2 (30 mL) and washed with 0.3 N HCl (2 x 20 mL) followed by brine (15 mL). Drying (MgSO4), filtration and concentration afforded the crude product. Flash chromatography using a gradient (25 to 50%) of acetone in EtOAc as eluent afforded the title compound. Example 16 Preparation of L-prolinamide, N-(4-methoxy-1,4-dioxobutyl)-L-alanyl-L-alanyl-N-[2-(1-oxoproxy)-3-methoxy-1-(1-methylethyl)-3-oxo-1-propenyl]-, (E)- (SEQ. ID NO: 9)

Scheme A: MeOSuc-Ala-Ala-Pro-Val-CO₂CH₃ (103 mg, 0.20 mmol) is treated with propionic anhydride (0.26 mL, 2.0 mmol) in pyridine (1.0 mL) followed by preparative TLC in a manner analogous to that described in Example 1 to afford the title compound. Example 17 Preparation of L-prolinamide, N-(4-methoxy-1,4-dioxobutyl)-L-alanyl-L-alanyl-N-[2-(2-methyl-1-oxopropoxy)-3-methoxy-1-(1-methylethyl)-3-oxo-1-propenyl]-, (EV (SEO. ID No. 10)

Scheme A: Treatment of MeOSuc-Ala-Ala-Pro-Val-CO2CH3 (103 mg, 0.20 mmol) with isobutyric anhydride (0.33 mL, 2.0 mmol) in pyridine (1.0 mL) followed by preparative TLC in a manner analogous to that described in Example 1 affords the title compound.

Preferred embodiments of the compounds of the present invention are best realized by the compounds of formulas I or IA, wherein R₁ is an isopropyl or n-propyl group, preferably an isopropyl group, R₂ is a (C₁-C₄)-alkyl radical or a phenyl group, preferably a (C₁-C₄)-alkyl radical, R₃ is a (C₁-C₄)-alkyl radical or a phenyl group, preferably a (C₁-C₄)-alkyl radical, P₂ is Pro, Pip, Aze, Pro(4-OBzl) or Gly, wherein the nitrogen atom of the α-amino group is substituted by a radical R, wherein R is a (C₁-C₄)alkyl radical, a phenyl, benzyl, cyclohexyl, cyclohexylmethyl, cyclooctyl, 2-bicyclo[2.2.1]hexyl, morpholinyl, piperidinyl, pyridinyl or tetrahydroquinolinyl group, preferably Pro, P₃ is Ile, Val or Ala, P₄ is Ala or is not present and K is an acetyl, t-butyloxycarbonyl, succinyl, methoxysuccinyl, 4-((chlorophenyl)sulfonylaminocarbonyl)phenylcarbonyl group or a radical of the formula

where Z is N or CH, B is a radical of the formulas

or

is.

Specific examples of preferred compounds include:

L-prolinamide, N-(4-methoxy-1,4-dioxobutyl)-L-alanyl-L-alanyl-N-[2-(acetyloxy)-3-methoxy-1-(1-methylethyl)-3-oxo -1-propenyl]-, (E)-; (SEQ. ID NO. 6)

L-prolinamide, N-(4-methoxy-1,4-dioxobutyl)-L-alanyl-L-alanyl-N-[2-(acetyloxy)-3-methoxy-1-(1-methylethyl)-3-oxo -1-propenyl]-, (Z)-; (SEQ. ID NO. 8)

L-prolinamide, N-(4-methoxy-1,4-dioxobutyl)-L-isoleucyl-N-[2-(acetyloxy)-3-methoxy-1-(1-methylethyl)-3-oxo-1-propenyl ]-, (E)-;

L-prolinamide, N-(4-methoxy-1,4-dioxobutyl)-L-valyl-N-[2-(acetyloxy)-3-methoxy-1-propyl-3-oxo-I-propenyl]-, ( E)-;

L-prolinamide, N-[4-[[[(4-chlorophenyl)sulfonyl]amino]carbonyl]benzoyl]-L-valyl-N-[2-(acetyloxy)-3-methoxy-1-(1-methylethyl) -3-oxo-I-propenyl]-, (E)-;

L-prolinamide, N-[4-[[[(4-chlorophenyl)sulfonyl]amino]carbonyl]benzoyl]-L-valyl-N-[2-(acetyloxy)-3-methoxy-I-(1-methylethyl) -3-oxo-1-propenyl]-, (Z)-;

L-prolinamide, N-[4-[[[(4-chlorophenyl)sulfonyl]amino]carbonyl]benzoyl]-L-valyl-N-[2-(acetyloxy)3-amino-1-(1-methylethyl)- oxo-1-propenyl]-, (E)-;

L-prolinamide, N-[4-[[[(4-chlorophenyl)sulfonyl]amino]carbonyl]benzoyl]-L-valyl-N-[2-(acetyloxy)-3-amino-1-(1-methylethyl) -3-oxo-1-propenyl]-, (Z)-;

L-prolinamide, N-[4-[[[(4-chlorophenyl)sulfonyl]amino]carbonyl]benzoyl]-L-valyl-N-[1-(1-methylethyl)-3-oxo-2-(acetyloxy) -3-(phenylamino)-1-propenyl]-, (E)-;

L-prolinamide, N-acetyl-L-valyl-N-[2-(acetyloxy)-3-ethoxy-1-(1-methylethyl)-3-oxo-1-propenyl]-, (E)-;

L-prolinamide, N-acetyl-L-valyl-N-[2-(acetyloxy)-3-ethoxy-I-(1-methylethyl)-3-oxo-1-propenyl]-, (Z)-;

L-prolinamide, N-acetyl-L-lysyl-N-[2-(acetyloxy)-3-ethoxy-I-(1-methylethyl)-3-oxo-1-propenyl]-, (E)-;

L-prolinamide, N-acetyl-L-prolyl-L-valyl-N-[2-(acetyloxy)-3-methoxy-1-(1-methylethyl)-3-oxo-I-propenyl]-, (E) -; (SEQ, ID NO. 7);

L-Prolinamide, N-[4-(4-Morpholinylcarbonyl)benzoyl]-L-valyl-N-[2-(acetyloxy)-3-ethoxy-1-(1-methylethyl)-3-oxo-1-propenyl] -, (E)-;

L-prolinamide, N-[4-(4-morpholinylcarbonyl]benzoyl]-L-valyl-N-[2-(acetyloxy)-3-ethoxy-1-(1-methylethyl)-3-oxo-1-propenyl] -, (Z)-;

L-prolinamide, N-(4-morpholinylcarbonyl)-L-valyl-N-[2-(acetyloxy)-3-ethoxy-1-(1-methylethyl)-3-oxo-1-propenyl]-, (E) -;

L-prolinamide, N-(4-morpholinylcarbonyl)-L-valyl-N-[2-(acetyloxy)-3-ethoxy-1-(1-methylethyl)-3-oxo-1-propenyl]-, (Z) -;

L-Prolinamide, N-(2-Furoyl)-L-valyl-N-[2-(acetyloxy)-3-ethoxy-1-(1-methylethyl)-3-oxo-1-propenyl]-, (E) -;

L-Prolinamide, N-[(Tetrahydro-2H-pyran-4-yl)carbonyl]-L-valyl-N-[2-(acetyloxy)-3-ethoxy-1-(1-methylethyl)-3-oxo- 1-propenyl]-, (E)-;

L-prolinamide, N-[3-(4-morpholinyl)-1,3-dioxopropyl]-L-valyl-N-[2-(acetyloxy)-3-ethoxy-1-(1-methylethyl)-3-oxo -I-propenyl]-, (E)-;

L-prolinamide, N-[3-(3-pyridyl)propanoyl]-L-valyl-N-[2-(acetyloxy)-3-ethoxy-1-(1-methylethyl)-3-oxo-1-propenyl] -, (E)-;

Glycinamide, N-(4-methoxy-1,4-dioxobutyl)-L-valyl-N-[2-(acetyloxy)-3-methoxy-1-(1-methylethyl)-3-oxo-I-propenyl-N² -(2,3-dihydro-1H-inden-2-yl)]-, (E)-;

Glycinamide, N-(4-methoxy-1,4-dioxobutyl)-L-valyl-N-[2-(acetyloxy)-3-methoxy-1-(1-methylethyl)-3-oxo-1-propenyl-N² -(2,3-dihydro-1H-inden-2-yl)]-, (Z)-;

Glycinamide, N-(4-methoxy-1,4-dioxobutyl)-L-valyl-N-[2-(acetyloxy)-3-methoxy-1-(1-methylethyl)-3-oxo-1-propenyl-N² -methyl]-, (E)-;

Glycinamide, N-(4-methoxy-1,4-dioxobutyl)-L-valyl-N-[2-(acetyloxy)-3-methoxy-1-(1-methylethyl)-3-oxo-1-propenyl-N² -methyl-, (Z)-;

Glycinamide, N-(4-methoxy-1,4-dioxobutyl)-L-valyl-N-[2-(acetyloxy)-3-methoxy-1-(1-methylefhyl)-3-oxo-1-propenyl-N² -cyclopentyl]-, (E)-;

Glycinannide, N-(4-methoxy-1,4-dioxobutyl)-L-valyl-N-[2-(acetyloxy)-3-methoxy-1-(1-methylethyl)-3-oxo-1-propenyl]- N²-cyclooctyl]-, (E)-;

Glycinamide, N-(4-methoxy-1,4-dioxobutyl)-L-valyl-N-[2-(acetyloxy)-3-methoxy-1-(1-methylethyl)-3-oxo-1-propenyl-N² -benzyl]-, (E)-;

L-prolinamide, N-(4-methoxy-1,4-dioxobutyl)-L-alanyl-L-alanyl-N-[2-(1-oxopropoxy)-3-methoxy-1-(1-methylethyl)-3 -oxo-1-propenyl]-, (E)-;

L-prolinamide, N-(4-methoxy-1,4-dioxobutyl)-L-alanyl-L-alanyl-N-[2-(2-methyl-1-oxopropoxy)-3-methoxy-1-(1- methylethyl)-3-oxo-1-propenyl]-, (E)-;

L-4-thiazolidamide, N-(4-methoxy-1,4-dioxobutyl)-L-valyl-N-[2-(acetyloxy)-3-methoxy-1-(1-methylethyl)-3-oxo-1 -propenyl]-, (E)-;

L-4-thiazolidamide, N-[1-(1-dimethylethoxy)carbony1]-L-valyl-N-[2-(acetyloxy)-3-methoxy-1-(1-methylethyl)-3-oxo-1- propenyl]-, (E)-;

L-1,2,3,4-Tetrahydro-3-isoquinolinamide, N-(4-methoxy-1,4-dioxobutyl)-L-valyl-N-[2-(acetyloxy)-3-methoxy-1-( 1-methylethyl)-3-oxo-1-propenyl]-, (E)- and

L-1,2,3,4-Tetrahydro-3-isoquinolinamide, N-[(1,1-dimethylethoxy)carbonyl]-L-valyl-N-[2-(acetyloxy)-3-methoxy-1-(1 -methylethyl)-3-oxo-1-propenyl]-, (E)-.

In another embodiment, the present invention provides a method of treating a patient suffering from a neutrophil-associated inflammatory disease comprising administering to the patient a therapeutically effective amount of a compound of formula I. The term "neutrophil-associated inflammatory disease" refers to diseases or conditions characterized by the migration of neutrophils to the site of inflammation and their participation in proteolytic degradation of basic biological substances. Neutrophil-associated associated inflammatory diseases for which treatment with a compound of formula I is particularly useful include: emphysema, cystic fibrosis, respiratory distress syndrome, septicemia, disseminated intravascular coagulation, gout, rheumatoid arthritis, chronic bronchitis and inflammatory bowel disease. Compounds of formula I which are particularly preferred for the treatment of neutrophil-associated inflammatory diseases include:

L-prolinamide, N-(4-methoxy-1,4-dioxobutyl)-L-alanyl-L-alanyl-N-[2-(acetyloxy)-3-methoxy-1-(1-methylethyl)-3-oxo -1-propenyl]-, (E)-; (SEQ. ID NO. 6)

L-prolinamide, N-(4-methoxy-1,4-dioxobutyl)-L-alanyl-L-alanyl-N-[2-(acetyloxy)-3-methoxy-1-(1-meihylethyl)-3-oxo -1-propenyl]-, (Z)-; (SEQ. ID NO. 8)

L-prolinamide, N-[4-[[[(4-chlorophenyl)sulfonyl]amino]carbonyl]benzoyl]-L-valyl-N-[2-(acetyloxy)-3-methoxy-1-(1-methylethyl) -3-oxo-1-propenyl]-, (E)- and

L-prolinamide, N-[4-[[[(4-chlorophenyl)sulfonyl]amino]carbonyl]benzoyl]-L-valyl-N-[2-(acetyloxy)-3-methoxy-1-(1-methylethyl) -3-oxo-1-propenyl]-, (Z)-.

As used here, the term "patient" refers to a warm-blooded animal, such as a mammal, suffering from a particular inflammatory disease state. It is understood that guinea pigs, dogs, cats, rats, mice, horses, cattle, sheep, and humans are examples of creatures within the scope of the meaning of the term.

The term "therapeutically effective amount" refers to an amount that, when administered as a single or multiple dose to a patient, is effective in alleviating symptoms associated with neutrophil-associated inflammatory diseases. As used herein, "relief of symptoms" of a respiratory disease refers to a reduction in severity compared to what would be expected without treatment and does not necessarily mean total elimination or cure of the disease. In determining the therapeutically effective amount or dose, a number of factors must be considered by the attending diagnostician, including, but not limited to: the type of mammal, its size, age and general health, the specific disease involved, the degree of involvement or severity of the disease, the response of the individual patient, the specific, administered compound, the mode of administration, the bioavailability characteristics of the administered product, the dose regimen selected, the use of concomitant medications, and other relevant circumstances.

A therapeutically effective amount of a compound of formula I is expected to vary from about 0.1 milligrams per kilogram of body weight per day (mg/kg/day) to about 100 mg/kg/day. Preferred amounts are expected to vary from 0.5 to about 10 mg/kg/day.

The compounds of this invention are prodrugs of a potent elastase inhibitor, particularly human neutrophil elastase, or are themselves elastase inhibitors. It is believed that the compounds of this invention exert their inhibitory activity by inhibiting the enzyme elastase, thereby providing relief from elastase-mediated diseases including, but not limited to: emphysema, cystic fibrosis, acute respiratory distress syndrome, septicemia, disseminated intravascular coagulation, gout, rheumatoid arthritis, chronic bronchitis, and inflammatory bowel disease. However, it is to be understood that the present invention is not limited to any particular theory or proposed mechanism for explaining efficacy in an end-use application.

In effectively treating a patient suffering from a disease state described above, a compound of formula I can be administered in any form or mode which renders the compound bioavailable in effective amounts, including oral, aerosol or parenteral routes. For example, compounds of formula I can be administered orally, by nebulization, subcutaneously, intramuscularly, intravenously, transdensally, intranasally, rectally, topically and the like. Oral or aerosol administration is generally preferred. One skilled in the art of preparing preparations can readily select the proper form and mode of administration depending on the particular characteristics of the compound chosen, the disease state to be treated, the stage of the disease and other relevant circumstances (Remington's Pharmaceutical Sciences, 18th Edition, Mack Publishing Co. (1990).

The compounds may be administered alone or in the form of a pharmaceutical composition together with pharmaceutically acceptable carriers or excipients, the ratio and type thereof being determined by the solubility and chemical properties of the compound selected, the route of administration chosen and standard pharmaceutical practice. The compounds of this invention, while effective in their own right, may be prepared in the form of their pharmaceutically acceptable salts, such as acid addition salts, for reasons of stability, ease of crystallization, higher solubility and the like.

In another embodiment, the present invention provides compositions comprising a compound of formula I in admixture or otherwise associated with one or more inert carriers. These compositions are useful, for example, as assay standards, as convenient means for bulk delivery, or as pharmaceuticals. An assayable amount of a compound of formula I is an amount that is readily measurable by standard assay procedures and techniques known and understood by those skilled in the art. Assessable amounts of a compound of formula I generally vary from about 0.001 to about 75% of the composition by weight. Inert carriers can be any material that does not decompose or otherwise covalently react with a compound of formula I. Examples of suitable inert carriers are water, aqueous buffers such as those generally useful in high performance liquid chromatography (HPLC) analysis, organic solvents such as acetonitrile, ethyl acetate, hexane and the like, and pharmaceutically acceptable carriers or excipients.

In particular, the present invention provides pharmaceutical compositions comprising a therapeutically effective amount of a compound of formula I in admixture or otherwise associated with one or more pharmaceutically acceptable carriers or excipients.

The pharmaceutical compositions are prepared in a manner known in the pharmaceutical art. The carrier or excipient may consist of a solid, semi-solid or liquid material which can serve as a vehicle or medium for the active ingredient. Suitable carriers or excipients are known in the art. The pharmaceutical composition may be adapted for oral, parenteral or topical use and may be administered to the patient in the form of of tablets, capsules, suppositories, solutions, suspensions or similar.

The compounds of the present invention can be administered orally, for example, with an inert diluent or with an edible carrier. They can be enclosed in gelatin capsules or compressed into tablet form. For the purpose of oral therapeutic administration, the compounds can be combined with excipients and used in the form of tablets, lozenges, capsules, elixirs, suspensions, syrups, wafers, chewing gums and the like. These preparations should contain at least 4% of the compound of the invention, the active ingredient, but this can be varied depending on the particular form and can conveniently be from 4% to about 70% of the weight of the unit. The amount of compound present in the compositions is such that a suitable dosage is obtained. Preferred compositions and preparations according to the present invention are prepared so that an oral dosage unit form contains between 5.0-300 milligrams of a compound of the invention.

The tablets, pills, capsules, lozenges and the like may also contain one or more of the following additives: binders such as microcrystalline cellulose, gum tragacanth or gelatin, excipients such as starch or lactose, disintegrants such as alginic acid, primogel, corn starch and the like, lubricants such as magnesium stearate or Sterotex, glidants such as colloidal silicon dioxide and sweeteners such as sucrose or saccharin may be added or a flavoring such as peppermint, methyl salicylate or orange flavor. If the dosage unit form is a capsule, it may contain, in addition to materials of the above type, a liquid carrier such as polyethylene glycol or a fatty oil. Other dosage unit forms may contain other various materials which modify the physical form of the dosage unit, e.g. as a coating agent. Thus, tablets or pills may be coated with sugar, shellac or other edible coating agents. A syrup may contain, in addition to the present compounds, sucrose as a sweetener and various preservatives, colorants and colouring and flavouring agents. Materials used in preparing these various compositions should be pharmaceutically pure and non-toxic in the amounts used.

For the purpose of parenteral therapeutic administration, the compounds of the present invention may be incorporated into a solution or suspension. These preparations should contain at least 0.1% of a compound of the invention, but this may be varied between 0.1 and about 50% by weight thereof. The amount of the compound of the invention present in such compositions is such that a suitable dosage is obtained. Preferred compositions and preparations according to the present invention are prepared so that a parenteral dosage unit contains between 5.0 to 100 milligrams of the compound of the invention.

The compounds of formula I of the present invention may also be administered by aerosol. The term "aerosol" is used to refer to a variety of systems ranging from colloidal in nature to pressurized pack systems. Delivery may be by a liquid or pressurized gas or a suitable pumping system which dispenses the active ingredients. Aerosols of the compound of formula I may be delivered in single-phase, biphasic or triphasic systems to deliver the active ingredient. The delivery system of the aerosol includes the necessary container, activators, valves, subcontainers and the like. Preferred aerosols may be determined by one of skill in the art.

The compounds of formula I of this invention may also be administered topically, and then the carrier may suitably comprise a solution, ointment or gel base. The base may comprise, for example, one or more of the following: petrolatum, lanolin, polyethylene glycols, beeswax, mineral oil, diluents such as water and alcohol, and emulsifiers and stabilizers. Topical preparations may contain a concentration of the compound of formula I or its pharmaceutically acceptable salt of from about 0.1 to about 10% w/v (weight per unit volume).

Some suitable transdermal devices are described in US Patent Nos. 3,742,951, 3,797,494, 3,996,934 and 4,031,894. These devices generally include a base material defining one of their surfaces, an adhesive layer permeable to the active agent defining the other surface and at least one reservoir containing the active agent interposed between the surfaces. In another In one embodiment, the active ingredient may be contained in a plurality of microcapsules which are distributed through the permeable adhesive layer. In each case, the active ingredient is evenly released from the reservoir or microcapsules through a membrane into the active ingredient-permeable adhesive layer which is in contact with the skin or mucous membrane of the recipient. When the active ingredient is absorbed through the skin, a controlled, predetermined flow of the active ingredient is administered to the recipient. In the case of microcapsules, the encapsulating agent may also act as a membrane.

In another device for transdermally administering the compounds according to the present invention, the pharmaceutically active compound is contained in a matrix from which it is released at the desired gradual, constant and controlled rate. The matrix is permeable to release of the compound by diffusion or microporous flow. The rate of release is controlled. Such a system, which does not require a membrane, is described in U.S. Pat. No. 3,921,636. At least two types of release are possible in these systems. Release by diffusion occurs when the matrix is nonporous. The pharmaceutically active compound dissolves in and diffuses through the matrix itself. Release by microporous flow occurs when the pharmaceutically active compound is transported through a liquid phase in the pores of the matrix.

The solutions or suspensions may also include one or more of the following additives: sterile diluents such as water for injection, saline, fixed oils, polyethylene glycols, glycerin, propylene glycol or other synthetic solvents, antibacterial agents such as benzyl alcohol or methylparaben, antioxidants such as ascorbic acid or sodium bisulfite, complexing agents such as ethylenediaminetetraacetic acid, buffers such as acetates, citrates or phosphates and tonicity adjusters such as sodium chloride or dextrose. The parenteral preparation may be enclosed in ampoules, single-use syringes or multidose vials made of glass or plastic.

The compounds of formula I are converted in vivo by esterases into compounds which are known to be effective as elastase inhibitors in humans. For example, compounds of formula I are converted into compounds disclosed in European Patent Application OPI No. 0195212, published September 24, 1986 and in Peet, NP et al., J. Med. Chem. 33 (1990), 394-407, which documents are incorporated herein by reference as fully set forth.

The efficacy of the compounds to inhibit elastase or to act as prodrugs of elastase inhibitors and the utility of the compounds of formulas I and IA in the treatment of neutrophil-associated inflammatory diseases can be demonstrated by recognized and reliable in vitro and in vivo models.

Example 18 In vitro test of elastase in the presence of MDL 104,569 and pig liver esterase

Elastase was assayed in vitro using the commercially available chromogenic substrate N-MeOSUc-Ala-Ala-Pro-Val-p-nitroanilide ([S] = 0.20 mM, Km = 0.16 mM). Assay techniques are similar to those described by Mehdi et al., Biochemical and Biophysical Research Communications 166 (1990), 595. The test mixture consists of partially purified elastase and substrate (0.2 mM) in 0.1 M HEPES (pH 7.5), 0.5 M NaCl, 10% DMSO and 0.1% Brij 35. The reaction mixture (1 or 2 mL in a plastic cuvette) is kept at 37ºC and hydrolysis of the substrate is followed in the presence of MDL 104,569 and 5 units/mL of porcine liver esterase (Sigma Chemical Co., Cat. No. E-3128). The elastase was isolated from human sputum, although it has recently become commercially available. The rate of hydrolysis of the substrate without added inhibitor or precursor is reported as 100%. In the presence of 1 µM MDL 104,569 a rate of 81% is obtained; When esterase (Leber-Procin, Sigma Chemical Co.) was also present, the rate decreased to 24%. At a concentration of 10 μM, the rate observed from the final level of inhibition in the assay in the presence of esterase was 130 nM, compared to a Ki of 200 nM for the independently measured starting level.

Example 19 In vitro test of elastase in the presence of MDL 105.565 and pig liver esterase

Elastase was tested in vitro in the presence of MDL 105,565 using the techniques and procedures described in Example 18. With MDL 105,565, a rate of 99% was obtained at a concentration of 10 nM; when esterase was added, this rate was 76%. At a concentration of 45 nM, the rates were 94% (without esterase) and 37% (with esterase). In this case, the Ki of the drug released was calculated to be 13 nM, compared to an independently determined Ki of 2 nM.

Example 20 HNE-induced pulmonary hemorrhage in hamsters

Acute lung injury induced by HNE was measured using a pulmonary hemorrhage model as described, e.g., by Fletcher, DS et al., Am. Rev. Respir. Dis. 141 (1990), 672-677, Skiles, JW et al., J. Med. Chem. 35 (1992), 641-662, Shah, SK et al., J. Med. Chem 35 (1992), 3745-3754, or Durham, SL et al., J. Pharm. Exp. Ther. 270 (1994), 185-141. HNE (10-25 µg/hamster in 0.05 M sodium acetate buffered saline) is administered as previously described by Schranfnagel, D. et al., Am. Rev. Resp. Dis. 129 (1984), A324, is instilled into CO2-anesthetized male Golden Syrian hamsters weighing 75 to 125 g (Charles River, NY). Instillation is performed using a blunt 20-gauge, 3-inch stainless steel needle inserted into the trachea at a point anterior to the carina using a fiber optic light source positioned at the animal's larynx. The volume of HNE or vehicle is approximately 100 µl. Animals are euthanized by CO2 asphyxiation 1 hour later and exsanguinated by transection of the inferior vena cava below the liver. The trachea is exposed and cannulated with PE-100 tubing (Clay Adams, Parsippany, NJ) and the BAL fluid is collected by gently instilling and withdrawing a single volume of saline (0.04 ml saline/g) three times. The hemoglobin content of the BAL Fluid is determined using a spectrophotometric test procedure and the amount is determined with reference to a Hgb standard curve. The results are expressed in milligrams per milliliter per Hgb ± SE.

Inhibition of HNE-induced pulmonary hemorrhage is determined by administering a dose of a compound of formulas I or IA to the hamster p.o., i.v., or i.t. before or after i.t. instillation of the HNE. The vehicles for administering a dose of the compounds of formulas I or IA to animals consist of 20% emulphor/water (p.o.), 0.2% triethylamine/saline (i.v.), and 10% dimethyl sulfoxide (i.t.). Appropriate vehicle controls are included in all experiments. Hamsters are administered oral doses of drug or vehicle (5 ml/kg) using a 20-gauge curved needle attached to a 1 ml syringe (5 ml/kg). Intravenous injections (2 mlg) are performed via the jugular vein using a 26 gauge ½ inch needle attached to a 1 ml syringe. Administration of the drug and vehicle into the trachea is as described above for HNE instillation.

SEQUENCE LISTING

(1) GENERALITIES:

(i) APPLICANT:

(A) NAME: Hoechst Marion Roussel, Inc.

(B) STREET: 2110 E. Gaibraith Rd., P.O. Box 156300

(C) CITY: Pittsburgh

(D) STATE: Ohio

(E) COUNTRY: USA

(F) POSTAL CODE (ZIP): 45215-6300

(G) TELEPHONE: 513-948-6566

(H) TELEFAX: SI3-948-7961 or 4681

(I) TELEX: 214320

(ii) TITLE OF INVENTION: Acylated Enol Derivatives of alpha-Xetoesters

and alpha-ketoamides

(iii) NUMBER OF SEQUENCES: 10

(iv) COMPUTER-READABLE FORM:

(A) MEDIUM TYPE: Floppy disk

(B) COMPUTER: IBM PC compatible

(C) OPERATING SYSTEM: PC-DOS/MS-DOS

(D) SOFTWARE: Patent Release #1.0. Version #1.30 (EPO)

(vi) PRIOR APPLICATION DATA:

(A) APPLICATION NUMBER: US 08/566,196

(B) FILING DATE: OI-DEC-1995

(2) INFORMATION FOR SEQ ID NO: 1:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 4 amino acids

(B) TYPE: amino acid

(C) STRANDEDNESS:

(D) TOPOLOGY: linear

(ii) MOLECULE TYPE: oeatide (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 1:

(2) INFORMATION FOR SEQ ID NO: 2:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 4 amino acids

(3) TYPE: amino acid

(C) STRANDEDNESS:

(D) TOPOLOGY: linear

(ii) MOLECULE TYPE: peptide (xi) SEQUENCE DESCRIPTION: SEQ iD NO: 2:

(2) INFORMATION FOR SEQ ID NO: 3:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 4 amino acids

(B) TYPE: amino acid

(C) STRANDEDNESS:

(D) TOPOLOGY: linear

(ii) MOLECULE TYPE: peptide (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 3:

(2) INFORMATION FOR SEQ ID NO: 4:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 4 amino acids

(B) TYPE: amino acid

(C) STRANDEDNESS:

(D) TOPOLOGY: linear

(ii) MOLECULE TYPE: peptide (xi) SEQUENCE DESCPRIPTION: SEQ ID NO: 4:

(2) INFORMATION FOR SEQ ID NO: 5:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 4 amino acids

(B) TYPE: amino acid

(C) STRANDEDNESS:

(D) TOPOLOGY: linear

(ii) MOLECULE TYPE: peptide (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 5:

(2) INFORMATION FOR SEQ ID NO: 6:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 4 amino acids

(B) TYPE: amino acid

(C) STRANDEDNESS:

(D) TOPOLOGY: linear

(ii) MOLECULE TYPE: peptide (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 6:

(2) INFORMATION FOR SEQ ID NO: 7:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 4 amino acids

(B) TYPE: amino acid

(C) STRANDEDNESS:

(D) TOPOLOGY: linear

(ii) MOLECULE TYPE: peptide (xi) SEQUENOS DESCRIPTION: SEQ ID NO: 7:

(2) INFORMATION FOR SEQ ID NO: 8:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 4 amino acids

(H) TYPE: amino acid

(C) STRANDEDNESS:

(D) TOPOLOGY: linear

(ii) MOLECULE TYPE: peptide (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 8:

(2) INFORMATION FOR SEQ ID NO: 9:

(i) SEQUENCE CERATIC ACTERISTICS:

(A) LENGTH: 4 amino acids

(B) TYPE: amino acid

(C) STRANDEDNESS:

(D) TOPOLOGY: linear

(ii) MOLECULE TYPE: peptide (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 9:

(2) INFORMATION FOR SEQ ID NO: 10

(i) SEQUENCE CERATIC ACTERISTICS:

(A) LENGTH: 4 amino acids

(B) TYPE: amino acid

(C) STRANDEDNESS:

(D) TOPOLOGY: linear

(ii) MOLECULE TYPE: peptide (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 10:

Claims (17)

1. Connecting the formulas or where R₁ is a (C₁-C₄)alkyl group, R₂ is a (C₁-C₄)alkyl group, a phenyl, benzyl, cyclohexyl or cyclohexylmethyl group, X is -CO₂R₃ or -CONHR₃', where R₃ is a (C₁-C₄)alkyl radical, a phenyl, benzyl, cyclohexyl or cyclohexylmethyl group and R₃' is a hydrogen atom, a (C₁-C₄)alkyl radical, a phenyl, benzyl, cyclohexyl or cyclohexylmethyl group, P₂ is Gly or Ala, wherein the nitrogen of the α-amino group is optionally substituted by a radical R, wherein R is (C₁-C₆)alkyl, (C₃-C₁₁)cycloalkyl, (C₃-C₁₂)cycloalkyl-(C₁-C₆)alkyl, (C₄-C₁₁)bicycloalkyl, (C₄-C₁₁)bicycloalkyl-(C₁-C₆)alkyl, (C₆-C₁₁)aryl, (C₆-C₁₀)aryl-(C₁-C₆)alkyl, (C3 -C7 )heterocycloalkyl, (C3 -C7 )heterocycloalkyl, (C1 -C6 )alkyl, (C5 -C9 )heteroaryl, (C5 -C9 )heteroaryl (C1 -C6 )alkyl , fused (C6 -C10 )-aryl-(C3 -C12 )cycloalkyl, fused (C6 -C10 )aryl-(C3 -C12 )cycloalkyl-(C1 -C6 )alkyl, is fused (C5 -C9 )-heteroaryl-(C3 -C12 )cycloalkyl or fused (C5 -C9 )heteroaryl-(C3 -C12 )cycloalkyl-(C1 -C6 )alkyl, or P&sub2; Pro, Aze, Ind, Tic, Pip, Tca, Pro(4-OBzl), Pro(4-OAc), Pro(4-OH) is, P₃ is Ala, bAla, Leu, Ile, Nle, Val, Nva, Lys or bVal, P₄ Ala, bAla, Val, Nva, bVal, Pro or absent, K is a hydrogen atom, an acetyl, succinyl, benzoyl, t-butyloxycarbonyl, carbobenzylvxy, dansyl, isovaleryl, -, methoxysuccinyl, 1-adamantanesulfonyl, 1-adamantaneacetyl, 2-carboxybenzoyl group, -C(O )N(CH3 )2 -, 4-((chlorophenyl)sulfonylaminocarbonyl)phenylcarbonyl, 4-((4-bromophenyl)sulfonylaminocarbonyl)phenylcarbonyl, 4-(sulfonylaminocarbonyl)phenylcarbonyl group or a residue of the formulas or where Z is N or CH, B is a radical of the formulas or is, R' is a hydrogen atom or a (C₁-C₄)alkyl radical, n is the number 0 or the integer 1 or 2, or a hydrate or a pharmaceutically acceptable salt thereof. 2. A compound according to claim 1, wherein R₁ is a methyl, propyl or isopropyl group, P₂ Gly or Ala, wherein the nitrogen atom of the α-amino group is optionally substituted by a radical R, wherein R is (C₁-C₆)alkyl, (C₃-C₁₂)cycloalkyl, cyclohexylmethyl, cyclopentylethyl, 2-bicyclo[1.1.0]butyl, 2-bicyclo[2.2.1]hexyl, 2-bicyclohexylmethyl, phenyl, 1-naphthyl, 2-naphthyl, benzyl, morpholinyl, piperidinyl, morpholinomethyl, pyridinyl, 2-quinoxalinyl, quinolinyl, 3-quinolinylmethyl, 2-indanyl or tetrahydroquinolinyl group, or P₂ Pro, Aze, Tic, Pip, Tca, Pro(OBzl), Pro(4-OAc) or Pro(4-OH), and P₃ is Ala, Leu, Ile, Nle, Val, Nva, or Lys and K is a hydrogen atom, an acetyl, succinyl, benzoyl, t-butyloxycarbonyl, carbobenzyloxy, dansyl, isovaleryl, methoxysuccinyl, 1-adamantanesulfonyl, 1-adamantanacetyl, 2-carboxybenzoyl group, -C(O)N(CH₃)₂, 4-((4-chlorophenyl)sulfonylaminocarbonyl)phenylcarbonyl, 4-((4-bromophenyl)sulfonylaminocarbonyl)phenylcarbonyl, 4-(sulfonylaminocarbonyl)phenylcarbonyl group or a radical of the formulas or where Z is N or CH, B is a radical of the formulas or is. 3. A compound according to claim 2, wherein X is -CO₂R₃. 4. A compound according to claim 2, wherein X is -CONHR₃. 5. A compound according to claim 3, wherein P₂ is Gly or Ala, the nitrogen atom of the α-amino group being optionally substituted by a radical R, wherein R is a methyl, cyclopentyl or 2-indanyl group or P₂ is Pro, Aze or Tic, P₃ is Ile, Val or Ala, P₄ is Ala, Pro or absent and K is an acetyl, succinyl, t-butyloxycarbonyl, carbobenzyloxy, methoxysuccinyl group, -C(O)N(CH₃)₂, 4-((4-chlorophenyl)sulfonylaminocarbonyl)phenylcarbonyl, 4-((4-bromophenyl)sulfonylaminocarbonyl)phenylcarbonyl, 4-(sulfonylaminocarbonyl)phenylcarbonyl group or a radical of the formula where Z is N or CH, B is a radical of the formulas or is. 6. A compound according to claim 4, wherein P₂ is Gly or Ala, the nitrogen atom of the α-amino group being optionally substituted by a radical R, wherein R is a methyl, cyclopentyl or 2-indanyl group or is P, Pro, Aze or Tic, P₃ is Ile, Val or Ala, P₄ is Ala, Pro or absent and K is an acetyl, succinyl, t-butyloxycarbonyl, carbobenzyloxy, methoxysuccinyl group, -C(O)N(CH₃)₂, 4-((4-chlorophenyl)sulfonylaminocarbonyl)phenylcarbonyl, 4-((4-bromophenyl)sulfonylaminocarbonyl)phenylcarbonyl, 4-(sulfonylaminocarbonyl)phenylcarbonyl group or a radical of the formula where Z is N or CH, B is a radical of the formulas or is. 7. A compound according to claim 5, wherein R1 is an isopropyl group, P2 is Pro and K is an acetyl, t-butyloxycarbonyl, succinyl, methoxysuccinyl, 4-((chlorophenyl)sulfonylaminocarbonyl)phenylcarbonyl group or a radical of the formula where Z is N or CH, B is a radical of the formulas or is. 8. A compound according to claim 6, wherein R₁ is an isopropyl group, P₂ is Pro and K is an acetyl, t-butyloxycarbonyl, succinyl, methoxysuccinyl, 4-((chlorophenyl)sulfonylaminocarbonyl)phenylcarbonyl group or a radical of the formula where Z is N or CH, B is a radical of the formulas or is. 9. A compound according to claim 1, wherein the compound is L-prolinamide, N-(4-methoxy- 1,4-dioxobutyl)-L-alanyl-L-alanyl-N-[2-(acetyloxy)-3-methoxy-1-(1-methylethyl)-3-oxo-1-propenyl]-, (E)-. 10. A compound according to claim 1, wherein the compound is L-prolinamide, N-[4-[[[(4- chlorophenyl)sulfonyl]amino]carbonyl]benzoyl]-L-valyl-N-[2-(acetyloxy)-3-methoxy-1-(1-methylethyl)-3-oxo-1-propenyl]-, (E)-. 11. A compound according to any one of claims 1 to 10 for use as a medicament. 12. A pharmaceutical composition comprising a compound according to any one of claims 1 to 10 and a pharmaceutically acceptable carrier. 13. Use of a compound according to any one of claims 1 to 10 for the manufacture of a medicament for inhibiting human neutrophil elastase in a patient in need thereof. 14. Use of a compound according to any one of claims 1 to 10 for preparing a medicament useful for treating a neutrophil-related inflammatory disease. 15. Use according to claim 14, wherein the neutrophil-associated inflammatory disease is emphysema. 16. Use according to claim 14, wherein the neutrophil-associated inflammatory disease is cystic fibrosis. 17. Use according to claim 14, wherein the neutrophil-associated inflammatory disease is chronic obstructive pulmonary disease.